A. A novel vibrational spectroscopic imaging system has been designed which combines the spectral resolving power of optical spectroscopy with the exquisite sensitivity and selectivity of vibrational infrared and Raman spectroscopies, techniques involving the absorption and inelastic scattering processes, respectively, of radiation by molecules. The imaging spectrometer, which is entirely solid state, is constructed around an acousto-optic tunable filter (AOTF) coupled to a refractive microscope optimized to the near-infrared or visible spectral regions with either a liquid nitrogen cooled, state-of-the-art, indium antinomide focal plane array detector or a silicon charge-coupled device detector. High fidelity Raman or near-infrared images are produced with unprecedented speeds; the maximum spatial resolution is about 1microm and is limited only by diffraction. The instrumentation accommodates both macroscopic and microscopic samples. The data generated by the spectrometer provides a series of 2-dimensional images as a function of wavelength. Since the data can also be viewed as a series of spatially resolved near-infrared or Raman spectra, the conformational and dynamic properties of specific domains within biological or chemical samples can be probed despite the existence of multiple components being present in complex matrices. A variety of model and intact membrane and polymer systems have been examined using either the near-infrared absorption, reflectance or Raman scattering modes of the imaging instrumentation. B. The oxidation-reduction properties of cytochrome oxidase and related systems have been examined by the simultaneous measurement of the resonance Raman and optical absorption spectra of potentiometrically controlled solutions. Redox data obtained from cytochrome oxidase in its native form argue against the traditional neoclassical models for the enzyme in which the heme a and a3 centers behave identically and titrate at the same voltages. Specifically, heme a displays effective midpoint potentials near 350 and 220 mV, while heme a3 exhibits lower Em's near 230 and 200 mV. For the cyanide bound complex of cytochrome oxidase, the heme a redox behavior is described by midpoint potentials at 350 and 260 mV. These data are consistent with the notion of anticooperative effects occurring with the heme and copper centers of the enzyme.